Chemokines are chemotactic cytokines, of molecular weight 6-15 kDa, that are released by a wide variety of cells to attract and activate, among other cell types, macrophages, T and B lymphocytes, eosinophils, basophils and neutrophils (reviewed in: Charo et al., New Eng. J. Med., 354:610-621 (2006); Luster, New Eng. J. Med., 338:436-445 (1998); and Rollins, Blood, 90:909-928 (1997)). There are two major classes of chemokines, CXC and CC, depending on whether the first two cysteines in the amino acid sequence are separated by a single amino acid (CXC) or are adjacent (CC). The CXC chemokines, such as interleukin-8 (IL-8), neutrophil-activating protein-2 (NAP-2) and melanoma growth stimulatory activity protein (MGSA) are chemotactic primarily for neutrophils and T lymphocytes, whereas the CC chemokines, such as RANTES, MIP-1α, MIP-1β, the monocyte chemotactic proteins (MCP-1, MCP-2, MCP-3, MCP-4, and MCP-5) and the eotaxins (-1 and -2) are chemotactic for, among other cell types, macrophages, T lymphocytes, eosinophils, dendritic cells, and basophils. There also exist the chemokines lymphotactin-1, lymphotactin-2 (both C chemokines), and fractalkine (a CX3C chemokine) that do not fall into either of the major chemokine subfamilies.
The chemokines bind to specific cell-surface receptors belonging to the family of G-protein-coupled seven-transmembrane-domain proteins (reviewed in: Horuk, Trends Pharm. Sci., 15:159-165 (1994)) which are termed “chemokine receptors.” On binding their cognate ligands, chemokine receptors transduce an intracellular signal though the associated trimeric G proteins, resulting in, among other responses, a rapid increase in intracellular calcium concentration, changes in cell shape, increased expression of cellular adhesion molecules, degranulation, and promotion of cell migration. There are at least ten human chemokine receptors that bind or respond to CC chemokines with the following characteristic patterns (reviewed in Zlotnik et al., Immunity, 12:121 (2000)): CCR-1 (or “CKR-1” or “CC-CKR-1”) [MIP-1α, MCP-3, MCP-4, RANTES] (Ben-Barruch et al., Cell, 72:415-425 (1993), and Luster, New Eng. J. Med., 338:436-445 (1998)); CCR-2A and CCR-2B (or “CKR-2A”/“CKR-2B” or “CC-CKR-2A”/“CC-CKR-2B”) [MCP-1, MCP-2, MCP-3, MCP-4, MCP-5] (Charo et al., Proc. Natl. Acad. Sci. USA, 91:2752-2756 (1994), and Luster, New Eng. J. Med., 338:436-445 (1998)); CCR-3 (or “CKR-3” or “CC-CKR-3”) [eotaxin-1, eotaxin-2, RANTES, MCP-3, MCP-4] (Combadiere et al., J. Biol. Chem., 270:16491-16494 (1995), and Luster, New Eng. J. Med., 338:436-445 (1998)); CCR-4 (or “CKR-4” or “CC-CKR-4”) [TARC, MDC] (Power et al., J. Biol. Chem., 270:19495-19500 (1995), and Luster, New Eng. J. Med., 338:436-445 (1998)); CCR-5 (or “CKR-5” OR “CC-CKR-5”) [MIP-1α, RANTES, MIP-1β] (Samson et al., Biochemistry, 35:3362-3367 (1996)); CCR-6 (or “CKR-6” or “CC-CKR-6”) [LARC] (Baba et al., J. Biol. Chem., 272:14893-14898 (1997)); CCR-7 (or “CKR-7” or “CC-CKR-7”) [ELC] (Yoshie et al., J. Leukoc. Biol., 62:634-644 (1997)); CCR-8 (or “CKR-8” or “CC-CKR-8”) [I-309] (Napolitano et al., J. Immunol., 157:2759-2763 (1996)); CCR-10 (or “CKR-10” or “CC-CKR-10”) [MCP-1, MCP-3] (Bonini et al., DNA Cell Biol., 16:1249-1256 (1997)); and CCR-11 [MCP-1, MCP-2, and MCP-4] (Schweickart et al., J. Biol. Chem., 275:9550 (2000)).
In addition to the mammalian chemokine receptors, mammalian cytomegaloviruses, herpesviruses and poxviruses have been shown to express, in infected cells, proteins with the binding properties of chemokine receptors (reviewed in: Wells et al., Curr. Opin. Biotech., 8:741-748 (1997)). Human CC chemokines, such as RANTES and MCP-3, can cause rapid mobilization of calcium via these virally encoded receptors. Receptor expression may be permissive for infection by allowing for the subversion of normal immune system surveillance and response to infection. Additionally, human chemokine receptors, such as CXCR4, CCR-2, CCR-3, CCR-5 and CCR-8, can act as co-receptors for the infection of mammalian cells by microbes as with, for example, the human immunodeficiency viruses (HIV).
The chemokines and their cognate receptors have been implicated as being important mediators of inflammatory, infectious, and immunoregulatory disorders and diseases, including asthma and allergic diseases; as well as autoimmune pathologies, such as rheumatoid arthritis and multiple sclerosis; and metabolic diseases, such as atherosclerosis and diabetes (reviewed in: Charo et al., New Eng. J. Med., 354:610-621 (2006); Gao, Z. et al., Chem. Rev., 103:3733 (2003); Carter, P. H., Curr. Opin. Chem. Biol., 6:510 (2002); Trivedi et al., Ann. Reports Med. Chem., 35:191 (2000); Saunders et al., Drug Disc. Today, 4:80 (1999); Premack et al., Nature Medicine, 2:1174 (1996)). For example, the chemokine monocyte chemoattractant-1 (MCP-1) and its receptor CC Chemokine Receptor 2 (CCR-2) play a pivotal role in attracting leukocytes to sites of inflammation and in subsequently activating these cells. When the chemokine MCP-1 binds to CCR-2, it induces a rapid increase in intracellular calcium concentration, increased expression of cellular adhesion molecules, and the promotion of leukocyte migration. Demonstration of the importance of the MCP-1/CCR-2 interaction has been provided by experiments with genetically modified mice. MCP-1−/− mice were unable to recruit monocytes into sites of inflammation after several different types of immune challenge (Lu, B. et al., J. Exp. Med., 187:601 (1998)). Likewise, CCR-2−/− mice were unable to recruit monocytes or produce interferon-γ when challenged with various exogenous agents; moreover, the leukocytes of CCR-2 null mice did not migrate in response to MCP-1 (Boring, L. et al., J. Clin. Invest., 100:2552 (1997)), thereby demonstrating the specificity of the MCP-1/CCR-2 interaction. Two other groups have independently reported equivalent results with different strains of CCR-2−/− mice (Kuziel, W. A. et al., Proc. Natl. Acad. Sci. USA, 94:12053 (1997), and Kurihara, T. et al., J. Exp. Med., 186:1757 (1997)). The viability and generally normal health of the MCP-1−/− and CCR-2−/− animals is noteworthy, in that disruption of the MCP-1/CCR-2 interaction does not induce physiological crisis. Taken together, these data lead one to the conclusion that molecules that block the actions of MCP-1/CCR-2 would be useful in treating a number of inflammatory and autoimmune disorders (reviewed in: Feria, M. et al., Exp. Opin. Ther. Patents, 16:49 (2006); and Dawson, J. et al., Exp. Opin. Ther. Targets, 7:35 (2003)). This hypothesis has now been validated in a number of different animal disease models, as described below.
It is known that MCP-1 is upregulated in patients with rheumatoid arthritis (Koch, A. et al., J. Clin. Invest., 90:772-779 (1992)). Moreover, several preclinical studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating rheumatoid arthritis. A DNA vaccine encoding MCP-1 was shown recently to ameliorate chronic polyadjuvant-induced arthritis in rats (Youssef, S. et al., J. Clin. Invest., 106:361 (2000)). Likewise, the disease symptoms could be controlled via direct administration of antibodies for MCP-1 to rats with collagen-induced arthritis (Ogata, H. et al., J. Pathol., 182:106 (1997)), or streptococcal cell wall-induced arthritis (Schimmer, R. C. et al., J. Immunol., 160:1466 (1998)). Perhaps most significantly, a peptide antagonist of MCP-1, MCP-1 (9-76), was shown both to prevent disease onset and to reduce disease symptoms (depending on the time of administration) in the MRL-1pr mouse model of arthritis (Gong, J.-H. et al., J. Exp. Med., 186:131 (1997)). Moreover, it has been demonstrated the administration of small molecule CCR-2 antagonists reduced clinical score in rodent models of arthritis (Brodmerkel, C. M. et al., J. Immunol., 175:5370 (2005); and Xia, M. et al., U.S. Publication No. 2006/0069123). Administration of an anti-CCR-2 antibody had varying effects on murine CIA, depending on the time of administration (Bruhl, H. et al., J. Immunol., 172:890 (2004)). Recent studies with CCR-2−/− mice have suggested that deletion of CCR-2 can exacerbate rodent arthritis models in specific experimental circumstances (Quinones, M. P. et al., J. Clin. Invest., 113:856 (2004); Quinones, M. P. et al., J. Mol. Med., 84:503 (2006)).
It is known that MCP-1 is upregulated in atherosclerotic lesions, and it has been shown that circulating levels of MCP-1 are reduced through treatment with therapeutic agents (Rezaie-Majd, A. et al., Arterioscler. Thromb. Vasc. Biol., 22:1194-1199 (2002)). Several key studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating atherosclerosis. For example, when MCP-1−/− mice are crossed with LDL receptor-deficient mice, an 83% reduction in aortic lipid deposition was observed (Gu, L. et al., Mol. Cell, 2:275 (1998)). Similarly, when MCP-1 was genetically ablated from mice which already overexpressed human apolipoprotein B, the resulting mice were protected from atherosclerotic lesion formation relative to the MCP-1+/+ apoB control mice (Gosling, J. et al., J. Clin. Invest., 103:773 (1999)). Likewise, when CCR-2−/− mice are crossed with apolipoprotein E−/− mice, a significant decrease in the incidence of atherosclerotic lesions was observed (Boring, L. et al., Nature, 394:894 (1998); Dawson, T. C. et al., Atherosclerosis, 143:205 (1999)). Finally, when apolipoprotein E−/− mice are administered a gene encoding a peptide antagonist of CCR-2, then lesion size is decreased and plaque stability is increased (Ni, W. et al., Circulation, 103:2096-2101 (2001)). Transplantation of bone marrow from CCR-2−/− mice into ApoE3-Leiden mice inhibited early atherogenesis (Guo, J. et al., Arterioscler. Thromb. Vasc. Biol., 23:447 (2003)), but had minimal effects on advanced lesions (Guo, J. et al., Arterioscler. Thromb. Vasc. Biol., 25:1014 (2005)).
Patients with type 2 diabetes mellitus typically exhibit insulin resistance as one of the hallmark features of the disease. Insulin resistance is also associated with the grouping of abnormalities known as the “metabolic syndrome” or “syndrome X,” which includes obesity, atherosclerosis, hypertension, and dyslipidemia (reviewed in: Eckel et al., Lancet, 365:1415 (2005)). It is well-recognized that inflammation plays a role in exacerbating the disease process in type 2 diabetes and the “syndrome X” pathologies (reviewed in: Chen, H., Pharmacological Research, 53:469 (2006); Neels et al., J. Clin. Invest., 116:33 (2006); Danadona et al., Am. J. Cardiol., 90:27 G-33G (2002); Pickup et al., Diabetologia, 41:1241 (1998)). MCP-1 is recognized as playing a role in obesity-induced insulin resistance. In culture, human preadipocytes constitutively expressed MCP-1 (Gerhardt, Mol. Cell. Endocrinology, 175:81 (2001)). CCR-2 is expressed on adipocytes; Addition of MCP-1 to differentiated adipocytes in vitro decreases insulin-stimulated glucose uptake and the expression of several adipogenic genes (LpL, adipsin, GLU-4), aP2, β3-adrenergic receptor, and PPARγ) (Sartipy, P. et al., Proc. Natl. Acad. Sci. USA, 96:6902 (1999)). Patients with type 2 diabetes had greater levels of circulating MCP-1 than non-diabetic controls (Nomura, S. et al., Clin. Exp. Immunol., 121:437 (2000)), and release of MCP-1 from adipose tissue could be reduced by treatment with anti-diabetic therapies such as metformin or thiazolidinediones (Bruun, J. M. et al., J. Clin. Endocrinol. Metab., 90:2282 (2005)). Likewise, MCP-1 was also overexpressed in murine experimental models of obesity, and was primarily produced by adipose tissue (Sartipy et al., Proc. Natl. Acad. Sci. USA, 100:7265 (2003)). In obese mice, the expression of MCP-1 both preceded and occurred concurrently with the onset of insulin resistance (Xu, H. et al., J. Clin. Invest., 112:1821 (2003)). Another study showed that the expression of MCP-1 positively correlated with body mass in the perigonadal adipose tissue of mice (Weisberg et al., J. Clin. Invest., 112:1796 (2003)). Consistent with these data, the development of insulin resistance in db/db mice was ameliorated either via genetic deletion of MCP-1 or by gene-induced expression of a dominant negative peptide (Kanda, H. et al., J. Clin. Invest., 116:1494 (2006)). The logical converse could also be demonstrated: overexpression of MCP-1 in adipose tissue promoted insulin resistance (Kamei, N. et al., J. Biol. Chem., 281:26602 (2006)). One conflicting result showing that genetic deletion of MCP-1 does not effect insulin resistance in the db/db mouse has also appeared (Chow, F. Y. et al., Diabetologia, 50:471 (2007)). Consistent with the data on MCP-1, direct studies with CCR-2 (the MCP-1 receptor) have showed that it plays a role in the formation of obesity and obesity-induced insulin resistance. Maintenance of a high fat diet increased the numbers of circulating CCR-2+ inflammatory monocytes in both wild-type (Tsou, C. L. et al., J. Clin. Invest., 117:902 (2007)) and ApoE−/− mice (Tacke, F. et al., J. Clin. Invest., 117:185 (2007)). Genetic deletion of CCR-2 reduced numbers of activated macrophages in murine adipose tissue (Lumeng, C. N. et al., Diabetes, 56:16 (2007)), but did not affect a population of M2 adipose macrophages thought to maintain the “lean” state (Lumeng, C. N. et al., J. Clin. Invest., 117:175 (2007)). Genetic deletion of CCR-2 reduced diet-induced obesity and improved insulin sensitivity in diet-induced obesity model (Weisberg, S. P. et al., J. Clin. Invest., 116:115 (2006); Cornelius, P. et al., PCT Publication No. WO 2006/013427 A2), depending on experimental conditions (Chen, A. et al., Obes. Res., 13:1311 (2005)). Administration of a small molecule CCR-2 antagonist also improved insulin sensitivity in this same model (Weisberg, S. P. et al., J. Clin. Invest., 116:115 (2006)).
Two studies described the important role of CCR-2 in hypertension-induced vascular inflammation, remodeling, and hypertrophy (Bush, E. et al., Hypertension, 36:360 (2000); Ishibashi, M. et al., Circ. Res., 94:1203 (2004)).
It is known that MCP-1 is upregulated in human multiple sclerosis, and it has been shown that effective therapy with interferon β-1b reduces MCP-1 expression in peripheral blood mononuclear cells, suggesting that MCP-1 plays a role in disease progression (Iarlori, C. et al., J. Neuroimmunol., 123:170-179 (2002)). Other studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating multiple sclerosis; all of these studies have been demonstrated in experimental autoimmune encephalomyelitis (EAE), the conventional animal model for multiple sclerosis. Administration of antibodies for MCP-1 to animals with EAE significantly diminished disease relapse (Kennedy, K. J. et al., J. Neuroimmunol., 92:98 (1998)). Furthermore, two reports have shown that CCR-2−/− mice are resistant to EAE (Fife, B. T. et al., J. Exp. Med., 192:899 (2000); Izikson, L. et al., J. Exp. Med., 192:1075 (2000)). A subsequent report extended these initial observations by examining the effects of CCR-2 deletion in mice from different strains (Gaupp, S. et al., Am. J. Pathol., 162:139 (2003)). Notably, administration of a small molecule CCR-2 antagonist also blunted disease progression in C57BL/6 mice (Brodmerkel, C. M. et al., J. Immunol., 175:5370 (2005)).
It is known that MCP-1 is upregulated in patients who develop bronchiolitis obliterans syndrome after lung transplantation (Reynaud-Gaubert, M. et al., J. Heart Lung Transplant., 21:721-730 (2002); Belperio, J. et al., J. Clin. Invest., 108:547-556 (2001)). In a murine model of bronchiolitis obliterans syndrome, administration of an antibody to MCP-1 led to attenuation of airway obliteration; likewise, CCR-2−/− mice were resistant to airway obliteration in this same model (Belperio, J. et al., J. Clin. Invest., 108:547-556 (2001)). These data suggest that antagonism of MCP-1/CCR-2 may be beneficial in treating rejection of organs following transplantation. In addition, studies have shown that disruption of MCP-1/CCR-2 axis was able to prolong the survival of islet transplant (Lee, I. et al., J. Immunol., 171:6929 (2003); Abdi, R. et al., J. Immunol., 172:767 (2004)). In rat graft models, CCR-2 and MCP-1 was shown to be upregulated in grafts that develop graft vasculopathy (Horiguchi, K. et al., J. Heart Lung Transplant., 21:1090 (2002)). In another study, anti-MCP-1 gene therapy attenuated graft vasculopathy (Saiura, A. et al., Arterioscler. Thromb. Vasc. Biol., 24:1886 (2004)). One study described inhibition of experimental vein graft neoinitimal formation by blockage of MCP-1 (Tatewaki, H. et al., J. Vasc. Surg., 45:1236 (2007)).
Other studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating asthma. Sequestration of MCP-1 with a neutralizing antibody in ovalbumin-challenged mice resulted in marked decrease in bronchial hyperresponsiveness and inflammation (Gonzalo, J.-A. et al., J. Exp. Med., 188:157 (1998)). It proved possible to reduce allergic airway inflammation in Schistosoma mansoni egg-challenged mice through the administration of antibodies for MCP-1 (Lukacs, N. W. et al., J. Immunol., 158:4398 (1997)). Consistent with this, MCP-1−/− mice displayed a reduced response to challenge with Schistosoma mansoni egg (Lu, B. et al., J. Exp. Med., 187:601 (1998)).
Other studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating kidney disease. Administration of antibodies for MCP-1 in a murine model of glomerularnephritis resulted in a marked decrease in glomerular crescent formation and deposition of type I collagen (Lloyd, C. M. et al., J. Exp. Med., 185:1371 (1997)). In addition, MCP-1−/− mice with induced nephrotoxic serum nephritis showed significantly less tubular damage than their MCP-1+/+ counterparts (Tesch, G. H. et al., J. Clin. Invest., 103:73 (1999)).
Several studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating systemic lupus erythematosus. CCR-2−/− mice exhibited prolonged survival and reduced renal disease relative to their WT counterparts in a murine model of systemic lupus erythematosus (Perez de Lema, G. et al., J. Am. Soc. Neph., 16:3592 (2005)). These data are consistent with the disease-modifying activity found in recent studies on genetic deletion of MCP-1 (Shimizu, S. et al., Rheumatology (Oxford), 43:1121 (2004); Tesch, G. H. et al., J. Exp. Med., 190:1813 (1999)) or administration of a peptide antagonist of CCR-2 (Hasegawa, H. et al., Arthritis Rheum., 48:2555 (2003)) in rodent models of lupus.
A remarkable 30-fold increase in CCR-2+ lamina propria lymphocytes was observed in the small bowels from Crohn's patients relative to non-diseased ileum (Connor, S. J. et al., Gut, 53:1287 (2004)). Also of note, there was an expansion in the subset of circulating CCR-2+/CD14+/CD56+ monocytes in patients with active Crohn's disease relative to controls. Several rodent studies have demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating Crohn's disease/colitis. CCR-2−/− mice were protected from the effects of dextran sodium sulfate-induced colitis (Andres, P. G. et al., J. Immunol., 164:6303 (2000)). Administration of a small molecule antagonist of CCR-2, CCR-5, and CXCR3 (murine binding affinities=24, 236, and 369 nM, respectively) also protected against dextran sodium sulfate-induced colitis (Tokuyama, H. et al., Int. Immunol., 17:1023 (2005)). Finally, MCP-1−/− mice showed substantially reduced colonic damage (both macroscopic and histological) in a hapten-induced model of colitis (Khan, W. I. et al., Am. J. Physiol. Gastrointest. Liver Physiol., 291:G803 (2006)).
Two reports described the overexpression of MCP-1 in the intestinal epithelial cells and bowel mucosa of patients with inflammatory bowel disease (Reinecker, H. C. et al., Gastroenterology, 108:40 (1995), and Grimm, M. C. et al., J. Leukoc. Biol., 59:804 (1996)).
One study described the association of promoter polymorphism in the MCP-1 gene with scleroderma (systemic sclerosis) (Karrer, S. et al., J. Invest. Dermatol., 124:92 (2005)). In related models of tissue fibrosis, inhibition of CCR-2/MCP-1 axis reduced fibrosis in skin (Yamamoto, T. et al., J. Invest. Dermatol., 121:510 (2003); Ferreira, A. M. et al., J. Invest. Dermatol., 126:1900 (2006)), lung (Okuma, T. et al., J. Pathol., 204:594 (2004); Gharaee-Kermani, M. et al., Cytokine, 24:266 (2003)), kidney (Kitagawa, K. et al., Am. J. Pathol., 165:237 (2004); Wada, T. et al., J. Am. Soc. Nephrol., 15:940 (2004)), heart (Hayashidani, S. et al., Circulation, 108:2134 (2003)), and liver (Tsuruta, S. et al., Int. J. Mol. Med., 14:837 (2004)).
One study has demonstrated the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating alveolitis. When rats with IgA immune complex lung injury were treated intravenously with antibodies raised against rat MCP-1 (JE), the symptoms of alveolitis were partially alleviated (Jones, M. L. et al., J. Immunol., 149:2147 (1992)).
Several studies have shown the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating cancer (reviewed in: Craig, M. J. et al., Cancer Metastasis Rev., 25:611 (2006); Conti, I. et al., Seminars in Cancer Biology, 14:149 (2004); Giles, R. et al., Curr. Cancer Drug Targets, 6:659 (2006)). When immunodeficient mice bearing human breast carcinoma cells were treated with an anti-MCP-1 antibody, inhibition of lung micrometastases and increases in survival were observed (Salcedo, R. et al., Blood, 96:34-40 (2000)). Using human clinical tumor specimens, CCR-2 expression was associated with prostrate cancer progression (Lu, Y. et al., J. Cell. Biochem., 101:676 (2007)). In vitro, MCP-1 expression has been shown to mediate prostrate cancer cell growth and invasion (Lu, Y. et al., Prostate, 66:1311 (2006)); furthermore, MCP-1 expressed by prostate cancer cells induced human bone marrow progenitors for bone resorption (Lu, Y. et al., Cancer Res., 67:3646 (2007)).
Multiple studies have described the potential therapeutic value of antagonism of the MCP-1/CCR-2 interaction in treating restenosis. In humans, MCP-1 levels correlate directly with risk for restenosis (Cipollone, F. et al., Arterioscler. Thromb. Vasc. Biol., 21:327 (2001)). Mice deficient in CCR-2 or in MCP-1 showed reductions in the intimal area and in the intima/media ratio (relative to wildtype littermates) after arterial injury (Roque, M. et al., Arterioscler. Thromb. Vasc. Biol., 22:554 (2002); Schober, A. et al., Circ. Res., 95:1125 (2004); Kim, W. J. et al., Biochem. Biophys. Res. Commun., 310:936 (2003)). In mice, transfection of a dominant negative inhibitor of MCP-1 in the skeletal muscle (Egashira, K. et al., Circ. Res., 90:1167 (2002)) also reduced intimal hyperplasia after arterial injury. Blockade of CCR-2 using a neutralizing antibody reduced neointimal hyperplasia after stenting in primates (Horvath, C. et al., Circ. Res., 90:488 (2002)).
Two reports describe the overexpression of MCP-1 rats with induced brain trauma (King, J. S. et al., J. Neuroimmunol., 56:127 (1994), and Berman, J. W. et al., J. Immunol., 156:3017 (1996)). In addition, studies have shown that both CCR-2−/− (Dimitrijevic, O. B. et al., Stroke, 38:1345 (2007)) and MCP-1−/− mice (Hughes, P. M. et al., J. Cereb. Blood Flow Metab., 22:308 (2002)) are partially protected from ischemia/reperfusion injury.
It is known that monocytes/macrophages play an important role in the development of neuropathic pain (Liu, T. et al., Pain, 86:25 (2000)). Consistent with this notion, a potential role for CCR-2 in the treatment of both inflammatory and neuropathic pain has been described recently. CCR-2−/− mice showed altered responses to inflammatory pain relative to their WT counterparts, including reduced pain behavior after intraplantar formalin injection and slightly reduced mechanical allodynia after intraplantar CFA injection (Abbadie, C. et al., Proc. Natl. Acad. Sci. USA, 100:7947 (2003)). In addition, CCR-2−/− mice did not display significant mechanical allodynia after sciatic nerve injury. Likewise, a small molecule CCR-2 antagonist reduced mechanical allodynia to ˜80% of pre-injury levels after oral administration (Abbadie, C. et al., PCT Publication No. WO 2004/110376).
One study described the critical role of MCP-1 in ischemic cardiomyopathy (Frangogiannis, N. G. et al., Circulation, 115:584 (2007)). Another study described the attenuation of experimental heart failure following inhibition of MCP-1 (Hayashidani, S. et al., Circulation, 108:2134 (2003)).
Other studies have provided evidence that MCP-1 is overexpressed in various disease states not mentioned above. These reports provide correlative evidence that MCP-1 antagonists could be useful therapeutics for such diseases. Another study has demonstrated the overexpression of MCP-1 in rodent cardiac allografts, suggesting a role for MCP-1 in the pathogenesis of transplant arteriosclerosis (Russell, M. E. et al., Proc. Natl. Acad. Sci. USA, 90:6086 (1993)). The overexpression of MCP-1 has been noted in the lung endothelial cells of patients with idiopathic pulmonary fibrosis (Antoniades, H. N. et al., Proc. Natl. Acad. Sci. USA, 89:5371 (1992)). Similarly, the overexpression of MCP-1 has been noted in the skin from patients with psoriasis (Deleuran, M. et al., J. Dermatol. Sci., 13:228 (1996), and Gillitzer, R. et al., J. Invest. Dermatol., 101:127 (1993)); correlative findings with predominance of CCR-2+ cells have also been reported (Vestergaard, C. et al., Acta Derm. Venerol., 84:353 (2004)). Finally, a recent report has shown that MCP-1 is overexpressed in the brains and cerebrospinal fluid of patients with HIV-1-associated dementia (Garzino-Demo, A., PCT Publication No. WO 99/46991).
In addition, CCR-2 polymorphism has been shown to be associated with sarcoidosis at least in one subset of patients (Spagnolo, P. et al., Am. J. Respir. Crit. Care Med., 168:1162 (2003)).
It should also be noted that CCR-2 has been implicated as a co-receptor for some strains of HIV (Doranz, B. J. et al., Cell, 85:1149 (1996)). It has also been determined that the use of CCR-2 as an HIV co-receptor can be correlated with disease progression (Connor, R. I. et al., J. Exp. Med., 185:621 (1997)). This finding is consistent with the recent finding that the presence of a CCR-2 mutant, CCR-2-64I, is positively correlated with delayed onset of HIV in the human population (Smith, M. W. et al., Science, 277:959 (1997)). Although MCP-1 has not been implicated in these processes, it may be that MCP-1 antagonists that act via binding to CCR-2 may have beneficial therapeutic effects in delaying the disease progression to AIDS in HIV-infected patients.
It should be noted that CCR-2 is also the receptor for the human chemokines MCP-2, MCP-3, and MCP-4 (Luster, New Eng. J. Med., 338:436-445 (1998)). Since the new compounds of formula (I) described herein antagonize MCP-1 by binding to the CCR-2 receptor, it may be that these compounds of formula (I) are also effective antagonists of the actions of MCP-2, MCP-3, and MCP-4 that are mediated by CCR-2. Accordingly, when reference is made herein to “antagonism of MCP-1,” it is to be assumed that this is equivalent to “antagonism of chemokine stimulation of CCR-2.”
Accordingly, compounds that modulate chemokine activity could demonstrate a wide range of utilities in treating inflammatory, allergic, autoimmune, metabolic, cancer and/or cardiovascular diseases. PCT Publication Nos. WO 2005/021500 A1 (incorporated herein by reference and assigned to present applicant), WO 2008/014381 A1, WO 2008/014360 A1 and WO 2008/014361 A1, disclose compounds that modulate MCP-1, MCP-2, MCP-3 and MCP-4 activity via CCR-2. The references also disclose various processes to prepare these compounds including multistep syntheses that include the introduction and subsequent removal of protecting groups.
It is desirable to find new compounds with improved pharmacological characteristics compared with known chemokine modulators. For example, it is desirable to find new compounds with equipotent dual CCR-2/5 inhibitory activity in comparison to selectivity for CCR-2 alone, predominantly CCR-2 versus CCR-5, predominantly CCR-5 versus CCR-2, or other G protein-coupled receptors (i.e., 5HT2A receptor). It is also desirable to find compounds with equipotent dual CCR-2/5 inhibitory activity and advantageous characteristics in one or more of the following categories:
(a) pharmaceutical properties (i.e., solubility, permeability, amenability to sustained release formulations);
(b) dosage requirements (e.g., lower dosages and/or once-daily dosing);
(c) factors which decrease blood concentration peak-to-trough characteristics (i.e., clearance and/or volume of distribution);
(d) factors that increase the concentration of active drug at the receptor (i.e., protein binding, volume of distribution);
(e) factors that decrease the liability for clinical drug-drug interactions (cytochrome P450 enzyme inhibition or induction, such as CYP 2D6 inhibition, see Dresser, G. K. et al., Clin. Pharmacokinet., 38:41-57 (2000), which is hereby incorporated by reference); and
(f) factors that decrease the potential for adverse side-effects (e.g., pharmacological selectivity beyond G protein-coupled receptors, potential chemical or metabolic reactivity, limited CNS penetration, ion-channel selectivity). It is especially desirable to find compounds having a desirable combination of the aforementioned pharmacological characteristics.
It is also desirable in the art to provide new and/or improved processes to prepare such compounds. These processes may be characterized, without limitation, by a) facile adaptation to larger scale production, such as pilot plant or manufacturing scales; b) process steps and/or techniques enabling improvements in the purity (including chiral purity), stability and/or ease of handling of intermediates and/or final compounds; and/or c) fewer process steps.